PROJECT SUMMARY A growing body of evidence points to cancer stem cells (CSCs) as the culprit behind persisting uncontrolled growth in several human malignancies, including one of the most lethal brain tumor Glioblastoma (GBM). It is hypothesized that CSCs in GBM (GSCs), with similar characteristics as normal tissue stem cells, are resistant to anti-cancer therapeutics and thus instrumental in initiating clinical relapse. Within the tumor specific ?niche?, a dynamic equilibrium exists between GSCs and lineage- committed cancer cells. This equilibrium is maintained by regulation of cell differentiation through a balance between asymmetric and symmetric cell division rates within the GSC compartment. This intrinsic homoeostatic state is critical for disease progression, as shifts in the equilibrium can influence the clinical outcome. For example, in the clinical setting GSC-rich tumors are more aggressive and associated with poor prognosis. It is, therefore, critical to elucidate the molecular mechanisms of how the stemness equilibrium state is maintained within the tumor microenvironment, as well as its contribution to therapeutic resistance and disease recurrence. To this end, we developed models for anti-glioma chemotherapy-induced recurrent GBM by using patient-derived xenograft (PDX), and investigated the evolutionary path to recurrence. Our data demonstrated that in the recurrence model, the equilibrium shifted toward a more stem-like state and therapy-induced interleukin 8 (IL8)-regulated epigenetic plasticity is critical for this change. Based on this, we hypothesize that cellular plasticity-mediated fate equilibrium shift towards a more stem-like state is responsible for the aggressiveness of recurrent GBMs and their resistance to conventional therapy. By using matched primary and recurrent patient GBM tissues as well as our PDX derived GBM models we now propose to investigate the role of therapy-induced cancer stem cell niche in disease recurrence (Aim 1), elucidate the molecular mechanisms stemness equilibrium within the tumor during and post therapy (Aim 2) and finally, examine the role of epigenetic plasticity in therapeutic resistance and disease recurrence (Aim 3). By elucidating the molecular mechanisms of intratumoral cell fate equilibrium and investigating how such equilibrium can be influenced by therapy will give us insight into their role in therapeutic resistance, disease progression as well as recurrence, and aid in identifying novel targets to prevent GBM recurrence.